Context: Recent studies of corticosteroid-binding globulin (CBG) indicate that it does not merely transport cortisol passively but also actively regulates its release in the circulation. We show how CBG binding affinity can vary to give changes in free cortisol concentration in a physiologically relevant range. Objective: The objective was to determine how the binding affinity of plasma CBG is affected by glycosylation, changes in body temperature, and the conformational change induced by proteases at sites of inflammation. Design: Binding assays were performed over a range of temperatures with plasma and recombinant CBG to determine the contribution of glycosylation. The role of conformational change was assessed by measuring binding affinities of plasma CBG before and after reactive loop cleavage by neutrophil elastase. Main Outcome Measures: Determination of binding constants allows calculation of clinically relevant changes in CBG saturation and free cortisol concentrations. Results: On reactive loop cleavage at inflammation sites, CBG can continue to act as a buffered source of cortisol, although with a much reduced affinity, to give a potential quadrupling of free cortisol. Predicted increases in systemic free cortisol resulting from elevated body temperatures, previously reported based on affinity measurements using nonglycosylated recombinant CBG, were shown here to be considerably increased using glycosylated plasma CBG, with a doubling for every 2°C rise in body temperature. Conclusions: The ability of CBG to modulate free cortisol levels in blood must be considered in the understanding and management of disease processes, as illustrated here with predictable changes in inflammation and fever.